NMR Structures of Proteins Using Stereospecific Assignments and Relaxation Matrix Refinement in a Hybrid Method of Distance Geometry and Simulated Annealing

  • J. Habazettl
  • M. Nilges
  • H. Oschkinat
  • A. T. Brünger
  • T. A. Holak
Part of the NATO ASI Series book series (NSSA, volume 225)


The hybrid method combining the early stages of a distance geometry program with molecular dynamics/simulated annealing in the presence of NMR constraints was optimized to obtain structures consistent with the observed NMR data. Two novel methods of stereospecific assignments of the protons at the prochiral carbons are used in simulated annealing, the “floating” chirality assignment and a high-dimensional potential. These two methods were compared with stereospecific assignments obtained from the coupling constant data. There is good agreement between the three methods in predicting the same stereospecific assignments. As the high-dimensional potential uses more relaxed absolute distance constraints and also takes into account the relative distance constraint patterns, it reduces possible overinterpretation of the NOE data. The structures obtained from the hybrid method were further refined using the relaxation matrix approach. This approach employs the analytical form of the gradient of the calculated spectrum. Compared to the structures determined with the two-spin approximation, the fit to the NMR data improves significantly with only minimal r.m.s. shifts in the structure during simple conjugate gradient minimization. The R-factors, defined similarly to the crystallographic R-factors, are 0.51 for the structures calculated using the two-spin approximation and 0.26 for the refined structures. Large shifts of approx. 1 Å occur during a dynamics/simulated annealing calculation. The various stages of refinement and stereospecific assignments are tested on the NOE data for the small squash trypsin inhibitor, CMTI-I. In the case of CMTI-I, the last step of the refinement improved the agreement with the X-ray structure significantly.


Target Function Cross Peak NOESY Spectrum Distance Geometry Relaxation Matrix 
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  1. 1.
    K. Wüthrich, in “NMR of Proteins and Nucleic Acids,” pp. 117–199, Wiley-Interscience, New York (1986).Google Scholar
  2. 2.
    R. Kaptein, E. R. P. Zuiderweg, R. M. Scheek, R. Boelens, and W. F. van Gunsteren, J. Mol. Biol. 182, 179–182 (1985).PubMedCrossRefGoogle Scholar
  3. 3.
    T. A. Holak, J. H. Prestegard, and J. D. Forman, Biochemistry 26, 4652–4660 (1987).PubMedCrossRefGoogle Scholar
  4. 4.
    T. A. Holak, S. K. Kearsley, Y. Kim, and J. H. Prestegard, Biochemistry 27, 6135–6142 (1988).PubMedCrossRefGoogle Scholar
  5. 5.
    R. Kaptein, R. Boelens, R. M. Scheek, and W. F. van Gunsteren, Biochemistry 27, 5389–5395 (1988).PubMedCrossRefGoogle Scholar
  6. 6.
    A. T. Brünger, G. M. Clore, A. M. Gronenborn, and M. Karplus, Proc. Natl. Acad. Sci. USA 83, 3801–3805 (1986).PubMedCrossRefGoogle Scholar
  7. 7.
    G. M. Clore and A. M. Gronenborn, C.R.C. in Biochemistry and Mol. Biol. 24, 479–564 (1989).CrossRefGoogle Scholar
  8. 8.
    M. Nilges, A. M. Gronenborn, A. T. Brünger, and G. M. Clore, Protein Eng. 2, 27–38 (1988).PubMedCrossRefGoogle Scholar
  9. 9.
    J. M. Moore, D. W. Case, W. J. Chazin, G. P. Gippert, T. F. Havel, R. Powls, and P. E. Wright, Science 240, 314–317 (1988).PubMedCrossRefGoogle Scholar
  10. 10.
    M. J. Tappin, A. Pastore, R. S. Norton, J. H. Freer, and I. D. Campbell, Biochemistry 27, 1643–1647 (1988).PubMedCrossRefGoogle Scholar
  11. 11.
    W. Braun and N. Gō, J. Mol. Biol. 186, 611–626 (1985).PubMedCrossRefGoogle Scholar
  12. 12.
    K. Wüthrich, Science 243, 45–50 (1989).PubMedCrossRefGoogle Scholar
  13. 13.
    W. Braun, Quart. Rev. Biophys. 19, 1115–1157 (1987).CrossRefGoogle Scholar
  14. 14.
    G. Wagner, W. Braun, T. F. Havel, T. Schaumann, N. Gö, K. Wüthrich, J. Mol. Biol. 196, 611–639 (1987).PubMedCrossRefGoogle Scholar
  15. 15.
    V. Saudek, R. J. P. Williams, and G. Ramponi, FEBS Lett. 242, 225–232 (1989).PubMedCrossRefGoogle Scholar
  16. 16.
    T. F. Havel, and K. Wüthrich, J. Mol. Biol. 182, 281–294 (1985).PubMedCrossRefGoogle Scholar
  17. 17.
    M. P. Williamson, T. F. Havel, and K. Wüthrich, J. Mol. Biol. 182, 295–315 (1985).PubMedCrossRefGoogle Scholar
  18. 18.
    T. A. Holak, M. Nilges, J. H. Prestegard, A. M. Gronenborn, and G. M. Clore, Eur. J. Biochem. 175, 9–15 (1988b).PubMedCrossRefGoogle Scholar
  19. 19.
    T. A. Holak, M. Nilges, and H. Oschkinat, FEBS Letters 242, 218–224 (1989).PubMedCrossRefGoogle Scholar
  20. 20.
    P. L. Weber, R. Morrison, and D. Hare, J. Mol. Biol. 204, 483–487 (1988).PubMedCrossRefGoogle Scholar
  21. 21.
    B. A. Borgias, M. Gochin, D. J. Kerwood, and T. L. James, Progress in NMR Spectr. 22, 83–100 (1990).CrossRefGoogle Scholar
  22. 22.
    R. Boelens, T. M. G. Koning, G. A. Van der Marel, J. H. van Boom, and R. Kaptein, R., J. Magn. Reson. 82, 290–308 (1989).Google Scholar
  23. 23.
    P. Yip, and D. A. Case, J. Magn. Reson. 83, 643–648 (1989).Google Scholar
  24. 24.
    A. T. Brünger, J. Mol. Biol. 203, 803–816 (1988).PubMedCrossRefGoogle Scholar
  25. 25.
    W. J. Metzler, D. R. Hare, and A. Pardi, Biochemistry 28, 7045–7052 (1989).PubMedCrossRefGoogle Scholar
  26. 26.
    T. F. Havel, I. D. Kuntz, and G. M. Crippen, Bull. Mth. Biol. 45, 673–698 (1983).Google Scholar
  27. 27.
    V. C. Singh, and P. A. Kollman, J. Comput. Chem. 5, 129–145 (1984).CrossRefGoogle Scholar
  28. 28.
    T. A. Holak, J. N. Scarsdale, and J. H. Prestegard, J. Magn. Reson. 74, 546–549 (1987).Google Scholar
  29. 29.
    K. Wüthrich, M. Billeter, and W. Braun, J. Mol. Biol. 169, 949–961 (1983).PubMedCrossRefGoogle Scholar
  30. 30.
    T. A. Holak, D. Gondol, J. Otlewski, and T. Wilusz, J. Mol. Biol. 210, 635–648 (1989).PubMedCrossRefGoogle Scholar
  31. 31.
    T. F. Havel, DISGEO, Quantum Chemistry Exchange, Program no. 507, Indiana University (1986).Google Scholar
  32. 32.
    J. Habazettl, C. Cieslar, H. Oschkinat, and T. A. Holak, FEBS Letters 268(1), 141–145 (1990).PubMedCrossRefGoogle Scholar
  33. 33.
    S. G. Hyberts, W. Mäki, and G. Wagner, Eur. J. Biochem. 164, 625–635 (1987).PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 1991

Authors and Affiliations

  • J. Habazettl
    • 1
  • M. Nilges
    • 2
  • H. Oschkinat
    • 1
  • A. T. Brünger
    • 2
  • T. A. Holak
    • 1
  1. 1.Max-Planck-Institut für BiochemieMartinsried bei MünchenGermany
  2. 2.Howard Hughes Medical Institute and Department of Molecular Biophysics and BiochemistryYale UniversityNew HavenUSA

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